Human chorionic gonadotrophin (hereinafter referred to as hCG) is composed of two subunits; the .alpha. subunit is largely identical to the .alpha. subunit of TSH, FSH and LH; the .beta. subunit also possesses areas which are homologous with these glycoprotein hormones as well as a 30 amino acid sequence that is not homologous. Many preparations of the .beta. subunit of hCG obtained by chromatography of urea dissociated hCG exhibit microheterogeneity, Domini et al, Acta Endocr. 73:133-145, 1973, i.e., they contain carbohydrate chains of different lengths and complexity. Besides other glcoprotein hormones or their metabolites, it is also possible that undisassociated hCG and subunit are present to some extent as contaminants in these preparations. These antigenic contaminants will thus also produce antibodies when injected along with the primary antigen in immunization procedures. It is therefore desirable to further purify the .beta.-hCG prior to antibody production in order to produce a highly specific antiserum with low cross reactivity to interfering substances. Such an antiserum would make possible the design of more sensitive agglutination reactions with antigen sensitized carriers.
It was thought that gel electrophoresis would provide a suitable approach to obtain the desired, purified antigen. Such a procedure for the separation of human serum proteins is described by Davis in Ann. N.Y. Ac. Sci., 121:404-427(1964) herein incorporated by reference. But, the staining procedure for localizing the separated protein requires prior fixation of the protein in acid which causes complete denaturation of the protein. Hartman et al in Anal. Biochem. 30:391-39(1969) herein incorporated by reference attempted to provide a method of staining the electrophoresed protein with minimal denaturation by utilizing anilinonaphthalene sulfonate to stain the gels once they are removed form their tubes following electrophoresis. It has been found, however, that this latter procedure is unsatisfactory with hCG and no staining is observed. The further suggestion by Hartman to utilize HCl to intensify the stain is also counterproductive causing further denaturation of the protein.